This invention relates to novel antibodies capable of specifically binding to the erbB-2, gp185.sup.erbB 2 gene product, and combinations thereof.
The invention further relates to the use of antibodies capable of binding to the erbB-2, and/or combinations thereof, for treating and/or preventing the formation of erbB-2 expressing tumors.
The invention also relates to the use of antibodies capable of binding to the erbB-2 gene product, and/or combinations thereof, for the immunological detection of erbB-2 expressing tumor cells in an analyte.
The invention further relates to conjugates comprising novel antibodies capable of binding to gp185.sup.erbB-2, and combinations thereof, wherein the antibody is bound to a cytotoxic moiety or a label which provides for the detection thereof, e.g., a fluorescent, enzymatic or radiolabel.
In particular, the invention relates to the use of monoclonal antibodies capable of specifically binding to gp185.sup.erbB-2, and/or combinations thereof, and/or single chain antibodies derived therefrom for the treatment and/or prevention of erbB-2 expressing tumors. Additionally, the invention relates to the use of specific monoclonal antibodies or single chain antibodies capable of specifically binding to gp185.sup.erbB-2 for the detection of erbB-2 expressing tumor cells.
In recent years, evidence has been accumulated that growth factors and their receptors may be involved in the process of malignant transformation. The erbB-2 gene product (also called HER2, neu or c-erbB-2) encodes a 185-kDa growth factor receptor which has been implicated in the malignancy of some human adenocarcinomas. Specifically, the erbB-2 protein, gp185.sup.erbB-2, is a receptor tyrosine kinase (Yarden et al, Ann. Rev. Biochem., (1988), 57, 443-478) which is homologous to the epidermal growth factor (EGF) receptor. (Coussens et al, Science, (1985), 230, 1132-1139; Yamamoto et al., Nature, (1986), 319, 230-234.) The rat homologue of the gene undergoes oncogenic activation through a single point mutation. (Bargmann et al, Cell, (1986), 45, 649-657.)
The erbB-2 protein, like other receptor proteins, is composed of extracellular, transmembrane and intracellular domains. The extracellular domain contains two cysteine-rich areas and is 44% homologous to the epidermal growth factor receptor (EGFR). The intracellular domain contains a tyrosine kinase which is 82% homologous to that of EGFR. Because of these similarities to EGFR and to other tyrosine kinase receptors, it has been suggested in the literature that the c-erbB-2 protein may function as a growth factor receptor.
Clinical and experimental evidence suggests a role for overexpression of the erbB-2 protein in the progression of human breast, ovarian, and non-small lung carcinoma. For example, amplification and/or overexpression of the erbB-2 gene have been shown in 20-30% of breast adenocarcinomas (King et al, Science, (1985), 229, 974-976; Slamon et al, Science, (1987), 235, 177-182; Slamon et al, Science, (1989), 244, 707-712; Yokota et al, Lancet, (1986), 1,765-767; King et al, Cancer Res., (1985), 49, 4185-4191), ovary adenocarcinomas (Slamon et al, Science, (1989), 244, 707-712), lung adenocarcinomas (Schneider et al, Cancer Res., (1989), 49, 4968-4971) and stomach adenocarcinomas (Park et al, Cancer Res., (1989), 49, 6605-6609). In breast carcinoma, a correlation has been observed between gene amplification and overexpression of erbB-2 protein and the aggressiveness of the malignancy (Slamon et al, Science, (1987), 237, 177-182; Slamon et al, Science, (1989), 244, 707-712). In cases of gene amplification, there is a resulting 50- to 100-fold increase in erbB-2 in RNA compared with normal cell levels (Kraus et al, EMBO J., (1987), 6, 605-610). The overexpression of erbB-2 has also been directly linked to the malignant conversion of cancer cells. (DiFiore et al, Science, (1986), 237, 178-182; Hudziak et al, Proc. Nat'l. Acad. Sci., (1987), 89, 7159-7163).
At least two lines of evidence suggest that erbB-2 overexpression may be involved in the pathogenesis of human neoplasia. First, as discussed supra, overexpression has been linked with poor prognosis in breast cancer, as well as ovarian cancer (Slamon et al, Science, (1989), 244, 707-712; Berchuk et al, Cancer Res., (1990), 50, 4087-4091), stomach cancer (Yonemura et al, Cancer Res., (1991), 51, 1004-1032) and lung cancer (Kern et al, Cancer Res., (1990), 50, 5184-5191). Second, artificial overexpression of erbB-2 induces a transformed phenotype in NIH 3T3 fibroblasts (DiFiore et al, Science, (1986), 237, 178-182; Hudziak et al, Proc. Nat'l. Acad. Sci., (1987), 84, 7159-7163), as well as in mammary epithelial cells (Pierce et al, Oncogene, (1991), 6, 1189-1194), suggesting that overexpression can not contribute directly to the development of the malignant phenotype.
Because of the extensive homology between the erbB-2 protein and the EGFR, it is widely assumed that the activation of growth signal transduction may proceed through similar mechanisms. One proposed mechanism involves receptor dimerization or oligomerization, which is thought to be an important step in the activation of EGFR intrinsic tyrosine kinase function (Yarden et al, Biochemistry, (1988), 27, 3114-3118; Schlessinger, Biochemistry, (1988), 27, 3119-3123). Interfering with receptor-receptor interactions has been evaluated as a potential therapeutic approach for the treatment of cancers associated with erbB-2 overexpression. In particular, such studies have involved the use of single monoclonal antibodies directed against the erbB-2 protein (Hudziak et al, Mol. Cell. Biol., (1989), 9, 1165-1172) and the related EGFR protein (Divgi et al, J. Nat'l. Cancer Inst., (1991), 83, 97-104) as potential therapeutic agents for the treatment of cancer.
The potential use of monoclonal antibodies for diagnosis and treatment of cancer has been studied extensively (Mellstadt, Curr. Opinion Immunol., (1990), 2, 708-713). Receptors for growth factors constitute a desirable target for this approach because their location on the cell membrane renders them accessible to antibody molecules. Moreover, antibodies directed to growth factor receptors can potentially block biological functions essential for cell proliferation.
Previous studies have demonstrated in animal systems the potential therapeutic effects of monoclonal antibodies against the EGFR (Matsui et al, Cancer Res., (1984), 44, 1002-1007; Aboud-Pirak et al, J. Nat'l. Cancer Inst., (1988), 80, 1605-1611). Also, different monoclonal antibodies to the erbB-2 receptor have been found to inhibit the proliferation of a human breast carcinoma cell line in human tissue culture (Hudziak et al, Mol. Cell. Biol., (1989), 9, 1165-1172), and an antibody directed to the rat erbB-2 protein, has been reported to inhibit the tumorigenicity of fibroblasts transformed by the mutant rat erbB-2 oncogene (Drebin et al, Proc. Nat'l. Acad. Sci., (1986), 83, 9126-9133; Drebin et al, Oncogene, (1988), 2, 387-399). Additionally, monoclonal antibodies which bind to erbB-2 have been used to study the biological function of the presumed receptor (McKenzie et al, Oncogene, (1989), 4, 543-548); Van Leenwen et al, Oncogene, (1990), 5,497,503; Fendly et al, Cancer Res. (1990), 50, 1550-1558).
While some monoclonal antibodies to the erbB-2 protein have shown promise as potential anticancer therapeutic agents, variable effects are observed dependent upon the particular monoclonal antibody. For example, Stancovski et al studied the in vivo effects of monoclonal antibodies on erbB-2 expressing tumors. Although some of the administered antibodies almost completely inhibited the growth in athymic mice of transfected murine fibroblasts that overexpress the erbB-2 protein, other antibodies were found to accelerate tumor growth or to result in intermediate responses (Stancovski et al, Proc. Nat'l. Acad. Sci., (1991), 88, 8691-8695). Based on the variable observed effects, Stancovski et al postulated that anti-erbB-2 antitumor antibodies may affect both receptor functioning and host-tumor interactions.
Recently, the construction and bacterial expression of a bifunctional single chain antibody-phosphatase fusion protein targeted to the human erbB-2 receptor was reported by Weis et al, Biotechnology, (1992), 10, 1128-1132. Wels et al, reported that this fusion protein exhibits sufficient binding affinity to cells expressing the erbB-2 receptor to permit the use thereof as an immunohistochemical reagent for the detection of erbB-2 antigen on cells or tissues.